FA gene mutation leads to MMP-7 overexpression and MAPK pathway activation. (A) TNF-α accumulation in the supernatant of AHH1 (normal), PD4L (FA-C), PD4L/FANCC (FA-C corrected), HSC72 (FA-A), and HSC72/FANCA (FA-A corrected) lymphoblasts. Supernatants were collected as described in Figure 1A. Data presented are the mean (± SD) of at least 3 independent experiments. *P < .01 compared with gene-corrected cells. (B) TNF-α accumulation in the supernatant of AHH1 (normal), HSC99 (FA-A), GM16757 (FA-F), and EUFA143 (FA-G) cell lines. Supernatants were collected as described in Figure 1A. Data presented are the mean (± SD) of at least 3 independent experiments. *P < .01. (C) HSC72, HSC99, PD4L, GM16757, and EUFA143 cells were treated with MMP inhibitor as described in Figure 3F and TNF-α content in supernatants was measured by ELISA. Data presented are the mean (± SD) of at least 3 independent experiments. *P < .01. (D) MMP-7 promoter activity in siRNA FANC–transfected HeLa cells. Cells were cotransfected with the reporter pGL3-hMMP-7 and phRL-TK, and induced luciferase activity was measured 24 hours later. Luciferase activity in GFP-siRNA–transfected cells was considered equal to 1 in each experiment. Data presented are the mean (± SD) of at least 3 independent experiments done in triplicate. *P < .01. (E) Western blot analysis of MAPK phosphorylation in siRNA FANC–transfected HeLa cells. Cells were collected 72 hours after transfection, and actin was used as a loading control.